PROJECT SUMMARY Sjgren's syndrome (SS) is a rheumatic autoimmune disease selectively targeting salivary and lacrimal glands, leading to painful dry mouth and eyes, oral infections, severe dental caries/tooth loss, fatigue, arthritis, neurologic involvement, and malignant B cell lymphoma. There are no effective biologic therapies. The strongest genetic risk factor is MHC class II DR3/DQ2. CD4+ T cells dominate salivary gland (SG) focal lymphocytic infiltrates, but B cells are also present. CD4+ T cells drive germinal center formation and class-switched, somatically hyper- mutated antibodies, all characteristic features of SS SG. Identification of the antigens and immunologic pathways driving the aberrant immune responses in SS is the focus of our proposal. In previous work, we isolated paired T cell receptor (TCR) ? and ? sequences of clonally expanded SG CD4+ T cells from DR3/DQ2+ patients and showed that the degree of clonal expansion correlated with reduced salivary flow and increased SG fibrosis. These are the first findings to link SG T cell infiltrates with low salivary flow and SG damage. Expanded clones within and between patients showed TCR amino acid similarities at sites contacting antigenic peptide, providing evidence that common antigen(s) drive SG CD4+ T cell proliferation. We used SG plasmablast-derived monoclonal antibodies (mAbs) from the same patients to identify 15 new candidate autoantigens. We also identified key transcripts overexpressed in SG CD4+ T cells that are characteristic of T follicular helper (Tfh) cells or may promote Tfh differentiation (TNFSF8-encoded CD30L). This proposal will test three hypotheses: i) previously unidentified antigens are targeted by SG CD4+ T cells and plasmablasts in SS, ii) autoreactivity to novel antigens may explain oral and neurologic manifestations of SS, and iii) clonally expanded SG CD4+ T cells express a unique transcriptional signature that promotes Tfh cell differentiation. Aim 1. Determine whether shared TCRs expressed on clonally expanded SG CD4+ T cells are specific for canonical and novel SS antigens bound by SG plasmablasts of the same subjects and are present in peripheral blood (PB) of SS cases, with frequencies reflecting SG disease features and severity. Aim 2. Identify the specificities of SS salivary gland plasmablast-derived mAbs and determine whether they have functional consequences and/or cross react with canonical SS antigens. Aim 3. Determine the transcriptional program of SG CD4+ T cells expressing shared, disease-associated TCRs, validate their predicted phenotypes and their localization in SG tissue, and test the hypothesis that CD30/CD30L signaling promotes human Tfh differentiation. This high impact research will identify autoantigens and SG T cell properties that may explain oral and neurologic SS pathology and that may be exploited to eliminate or regulate lymphocytes that drive chronic disease.